This is a competitive renewal of a program focused on exploring protein - carbohydrate-binding interactions using synthetic ligands. The research proposed has three objectives. First, we shall investigate the recognition properties of the receptors DC-SIGN and DC-SIGNR by synthesizing and identifying inhibitors of these lectins. DC-SIGN has been shown to mediate the interaction of dendritic cells with T cells. Both DCSIGN and DC-SIGNR can facilitate HIV infection of CD4-positive cells in trans. The physiological and pathological functions of DC-SIGN and DC-SIGNR are mediated through their interactions with N-linked carbohydrates. We propose to use methods developed in the past grant period to synthesize libraries of ligands and to identify active DC-SIGN and DC-SIGNR inhibitors. Both monovalent and multivalent inhibitors will be synthesized and tested for their ability to block key pathological and physiological functions of these lectins. Second, we shall use the inhibitors identified to probe the biological functions of DC-SIGN and DCSIGNR. Third, we shall continue our efforts to develop methods for the synthesis of N-linked glycopeptides. The proposed research is divided into 5 aims. The proposed studies in aim 1 are focused on developing a high throughput assay for identifying DC-SIGN and DC-SIGNR ligands. In Aim 2, we propose to synthesize and evaluate compounds libraries designed to inhibit DC-SIGN, DC-SIGNR and other C-type lectins. The goal of aim 3 is to generate multivalent compounds that inhibit and/or cluster DC-SIGN and DC-SIGNR. We postulate that multivalent ligands will more potently block the target lectins. Aim 4 is focused on testing the activities of synthetic inhibitors in relevant assays. The ligands for their abilities to bind to cells producing DC-SIGN or DC-SIGNR, to inhibit the binding of ICAM-3 to dendritic cells, to block the binding of HIV-1 gp120 to DC-SIGN(DC-SIGNR)-positive cells, to inhibit HIV transmission, and to promote DC-SIGN and DCSIGNR internalization. Aim 5 is focused on the development of a new approach to the convergent synthesis of glycopeptides. We anticipate that the results from our studies will provide new tools for the investigation of DC-SIGN and DC-SIGNR function in particular and for probing C-type lectin function in general.